Epigenetic and Expression Profiling of Tissue Selective Estrogen Compounds in Breast Epithelial Cell Lines (RNA-seq). Homo sapiens

Current approaches to menopausal hormone therapy consist of compounds which provide tissue specific beneficial effects while working to minimize any increase in cancer risk associated with replacement therapy. In this study, an examination of the genomic distribution of histone modification and expression changes in two breast epithelial cell lines, the normal MCF10A cell line and the MCF-7 breast cancer cell line, upon treatment with several of the current therapies used for treating post-menopausal symptoms is presented. Analysis of the observed changes upon treatment with either 17-estradiol (E2), a combined treatment with the top 10 most abundant Premarin chemical equivalents (EC10), the non-steroidal SERM, Bazadoxifene (BZA) and the TSEC; BZA combined with EC10 (Duavee) or BZA combined with E2 provides evidence of expression changes in genes involved in DNA replication, nucleosome assembly, RNA-splicing and processing, biological regulation and cell signaling, and many of the known steroid response genes. There were distinct global changes in chromatin states observed that were specific to each of the breast epithelial cell type normal, basal MCF10A and ESR/PGR positive luminal breast cancer MCF7 cell lines and selective changes observed upon treatment with each of the treatment groups. Specifically, we observed a global loss of H3K4ac upon treatment with BZA. These changes were observed in both ESR1 bound and non-bound regions suggesting modifications in both promoters and enhancer regions. The levels of H3K4ac upon treatment of BZA showed a decrease below basal non-treated H3K4ac levels in MCF7. There was an increase observed with E2 and EC10 treatment in MCF7 that was mitigated by the addition of BZA to either steroid as a combined treatment. This study provides a better understanding of the role of epigenetic modifications in endocrine responsiveness and provides evidence of an improved therapeutic profile for combined TSEC compounds for treating and managing the overall physiological effects of declining estrogen. Overall design: 6 drug treatments in 2 cell lines with 3 biological replicates

当前绝经激素治疗（menopausal hormone therapy）的策略多采用兼具组织特异性获益与降低替代治疗相关癌症风险升高效应的化合物。本研究针对两种乳腺上皮细胞系——正常基底型乳腺上皮细胞系MCF10A与雌激素受体1（ESR1）/孕激素受体（PGR）阳性的腔面型乳腺癌细胞系MCF-7——接受数种当前用于缓解绝经后症状的治疗方案后的组蛋白修饰基因组分布与基因表达变化展开分析。
本研究对以下处理组的细胞变化进行了分析：分别接受17β-雌二醇（17β-estradiol, E2）、由倍美力（Premarin）中丰度最高的10种化学等效物组成的联合制剂（EC10）、非甾体类选择性雌激素受体调节剂（selective estrogen receptor modulator, SERM）巴多昔芬（Bazadoxifene, BZA），以及组织选择性雌激素复合物（tissue-selective estrogen complex, TSEC）相关组合方案：即巴多昔芬联合EC10（商品名倍美罗，Duavee）、巴多昔芬联合E2。分析结果显示，DNA复制、核小体组装、RNA剪接与加工、生物学调控及细胞信号通路相关基因，以及众多已知的类固醇应答基因均出现表达变化。
研究观察到染色质状态出现显著的全局变化，且该变化呈现细胞类型特异性：正常基底型MCF10A细胞与ESR1/PGR阳性的腔面型乳腺癌MCF-7细胞各自具有独特的染色质调控特征；同时各给药处理组也呈现出特异性的细胞变化。具体而言，经巴多昔芬处理后，我们观察到全基因组范围内H3K4乙酰化修饰（H3K4ac）水平显著下降。该修饰变化同时出现在ESR1结合区域与非结合区域，提示启动子及增强子区域均发生了组蛋白修饰改变。在MCF-7细胞中，巴多昔芬处理后的H3K4ac水平降至未处理的基础水平以下。在MCF-7细胞中，17β-雌二醇与EC10处理可使H3K4ac水平升高，而将巴多昔芬与任一甾体类药物联合使用时，该升高效应均被削弱。
本研究进一步阐明了表观遗传修饰在内分泌应答中的作用，并为组织选择性雌激素复合物联合疗法改善绝经后雌激素水平下降所引发的整体生理效应提供了实验依据。
实验整体设计：两种细胞系各接受6种药物处理，每组设置3次生物学重复。